Further characterization of the binding of human recombinant interleukin 2 to heparin and identification of putative binding sites

We have previously provided compelling evidence that human recombinantinterleukin 2 (IL-2) binds to the sulfated polysaccharides heparin, highlysulfated heparan sulfate and fucoidan. Here we show that IL-2 binding isdependent on heparin chain length, but with fragments as small as 15-mersretaining binding activity. The addition of exogenous heparin has no effecton the in vitro biological activity of IL-2. In addition soluble IL-2receptor alpha and beta polypeptides do not compete with heparin for thebinding of IL-2. IL-2 bound by heparin is still recognized by two IL-2specific monoclonal antibodies, 3H9 and H2- 8, whose epitopes lie in theamino terminal region. Murine IL-2 unlike its human counterpart fails tobind to heparin. Human IL-2 analogs with single amino acid substitutions atpositions Lys43, Thr51, and Gln126 analogs no longer bind to heparin. Bycontrast the Arg38Ala analog retains heparin full heparin binding activity.These experimental findings together with molecular modeling studiessuggest two putative heparin binding sites on human IL-2, one involvingfour basic residues, Lys48, Lys49, Lys54, and His55, and the other being adiscontinuous site comprising Lys43, Lys64, Arg81, and Arg83. Neither ofthese two clusters is completely conserved in murine IL-2. Overall our datasuggest that the binding of human IL-2 to heparin and heparan sulfate doesnot interfere with IL-2/IL-2 receptor interactions. Therefore, binding toglycosaminoglycan may be a mechanism for retaining the cytokine in anactive form close to its site of secretion in the tissue, thus favoring aparacrine role for IL-2.